Identification of the first several nephrotic syndrome and focal segmental glomerulosclerosis (FSGS) genes has had a significant impact on the understanding of glomerular function and disease. However, the known genes account for only a fraction of these diseases. During the most recent period of this award, we identified mutations in INF2 as a relatively common cause of autosomal dominant FSGS. We also identified two coding variants in APOL1 that explain the high rate of FSGS and other non-diabetic kidney diseases in African Americans. Here our goal is further elucidation of the genetic basis of FSGS, using a database of over 3800 individuals assembled over the past sixteen years. Specifically: (1) We will perform mutational analysis of known FSGS genes in families with FSGS and in patients with sporadic FSGS. We will continue our ongoing ascertainment of families with FSGS as well as sporadic adult and pediatric cases. Mutational analysis of these known FSGS genes will further inform our understanding of the spectrum of mutations, inform genotype/phenotype relationships, provide further information regarding structure and function, and help clarify the utility of genetic testing. (2) We will also aim to identify new FSGS genes. We will use exome capture and next-generation sequencing in affected members of multiple families with inherited FSGS to in order to identify rare coding sequence variants. We will use a combination of known variant filtering, cosegregation analysis, and evolutionary conservation to define which of these genes and variants are likely causally related to the development of FSGS. We will perform replication studies in additional FSGS families and sporadic FSGS cases as well as functional assessments in order to prove causal relationships to disease. (3) We will also perform studies to identify de novo variants causing or contributing to FSGS using exome sequencing. We will use exome sequencing in sporadic FSGS cases and their parents in order to identify de novo variants that may cause or contribute to the development of FSGS. This is a potentially powerful approach because fewer than one de novo exomic variant is expected per individual, making the list of candidate genes expected from this approach very low. Based on the great recent success of this approach in studies of sporadic neuropsychiatric disorders, we anticipate that we will identify several genes in which de novo variants are seen in more than one affected individual, and/or specific pathways frequently mutated. We expect to see overlap among genes harboring de novo variants and genes in which inherited variants segregate with FSGS in families. FSGS is a significant and growing cause of chronic kidney disease and kidney failure. FSGS is also a common consequence of a variety of primary conditions. These studies will help understand the underlying causes of this disease.